p53 is a major determinant of tumor response to chemo/radiotherapy in part through regulation of cell cycle checkpoints and apoptosis. Recent evidence suggests that transcriptional activation by p53 is critical for apoptosis and tumor suppression. A number of p53 downstream target genes have been identified, including p21WAF1, KILLER/DR5, Fas/APO1, Bax, Bak, Noxa, PUMA, Bid, Caspase 6, and PIDD. Our recent studies have uncovered distinct patterns of p53 target gene transcriptional induction in vivo, both in magnitude of induction and location within tissue compartments, as well as a wide range of cell death induction in various tissues. We hypothesize these patterns dictate the phenotype of p53-induced death in vivo. We have also recently proposed a novel molecular mechanism of chemo/radiosensitivity that relies on the upregulation of a novel class of pro-apoptotic p53 targets which can lower the cell death threshold following exposure of cells to chemotherapy or radiation. Using microarray screens we have identified polo-like kinase 2 as a candidate p53 target gene and have found that RNA-interference with its expression can remarkably sensitize cancer cells to microtubule active chemotherapeutics. In the next funding period we propose to extend our findings through completion of the following specific aims: Aim 1: Identify and characterize candidate p53 target genes that regulate tissue radiosensitivity. Aim 2: Investigate the molecular basis of tissue and cell-specific p53 activation and p53 target gene regulation. Aim 3: Investigate the relative importance of specific p53 targets to irradiation-induced cell death of normal and tumor tissues using in vitro and in vivo strategies. Aim 4: Investigate the role of polo-like kinase 2 as a p53 target in control of a G2/M checkpoint and therapeutic response to radiation. Additional studies interacting with other projects include analysis of irradiation-induced foci (Tim Yen), analysis of defective apoptotic responses during metastases and p53 targets restricted in induction by cell cycle phase (Ruth Muschel) and analysis of pro-survival signals downstream of Ras and Akt in inhibition of radiosensitivity (Gillies McKenna and Eric Bernhard). Our studies are expected to further our understanding of the molecular basis of irradiation-induced apoptosis, the role of p53, and may lead to novel strategies for protection of tissues from toxicity and/or for enhanced tumor cell killing.